Electrophoretic gels, usually comprising hydrogels such as agarose or polyacrylamide are used for the separation of nucleic acids, proteins and other macromolecular compounds. The sample to be separated is placed at one end of the gel and a direct electric field is applied between the ends of the gel causing the components of the sample to migrate through the gel at rates dependent upon their molecular size and charge.
A mixture of components to be separated is normally introduced into one of a number of small wells formed in an upper edge of the gel before the electric current is applied. It is usual to run a number of such mixtures simultaneously on an electrophoretic gel in a side by side arrangement. For this purpose one mixture is placed in each of a series of wells formed in the upper edge of the gel.
In the past electrophoretic gels were formed by juxtaposing a pair of glass plates in a slightly spaced apart side-by-side relationship and filling the space therebetween with a liquid which can set and form an electrophoretic gel. The side edges of the space between the glass plates were typically sealed with adhesive tape or a similar material, and when the gel is poured a comb was placed in the upper end of the space between the glass plates. After the gel had set the comb would be withdrawn leaving a series of spaced apart wells defined in the top of the gel, each well having been defined by one tooth of the comb. A tongue of gel remains between the glass plates separating the pair of adjacent wells.
The gel is immersed in a buffer solution and electrodes above and below the gel cause a direct electric current to flow through the gel. The components in the mixtures travel through the gel from the top to the bottom of the gel at different rates depending on their size and charge and separate out into bands.
In more recent years it has been proposed to preform electrophoretic gels in cassettes formed of synthetic plastics materials. The side walls of the cassette are formed with integral means to connect them together along the sides of the cassette. Again when the gel is poured a comb is placed in the upper end of the space between the walls of the cassette for the formation of the spaced apart wells. However, this arrangement has a disadvantage that upon withdrawal of the comb the fingers of the gel may, with time, show an increased tendency to break away from the remainder of the gel. This results in poorly defined wells. Alternatively, if the tongues of gels are left intact upon withdrawal of the comb they may not firmly adhere to the plastic side wall of the cassette. This has resulted that the tongues may fall over side ways occluding an adjacent well.
One attempt to overcome this problem is described in U.S. Pat. No. 5,288,465, where ribs are provided in the cassette walls to define wells at one end of the cassette. Whilst this arrangement provides stable wells suitable to hold the sample material, this arrangement has a disadvantage that the solid walls of the wells interfere with the smooth flow of electric current through the electrophoretic gel: because of the ribs, the current is initially confined and then spreads out which has a result that as the run proceeds bands from adjoining wells tend to spread out and merge with one another.
International Patent Application No WO 97/04307 addresses the problem of current interference by replacing the ribs by a plurality of small projections or pegs. These projections or pegs extend into the gel to support the gel fingers. The projections support the gel whilst allowing substantially parallel flow of the current to establish before the current meets the samples positioned in the bottom of the wells. This parallel current flow is necessary to maintain the separation between the bands/samples from the different wells and prevent the samples for spreading outwards.
However, the disadvantage of the cassette shown in WO 97/04307 is that as the combs are withdrawn, the tops of the gel fingers tend to break away from the remainder of the gel finger.
The present invention is directed to alternative arrangements addressing the problems associated with spreading current and also the integrity of the walls of the sample wells.